U937 apoptotic cell death by nitric oxide: Bcl-2 downregulation and caspase activation.
Upon treatment with NO-releasing compounds such as S-nitrosoglutathione or spermine NO, human myeloid leukemia U937 cells undergo apoptosis. Early NO-mediated signals comprise activation of a Z-A-DCB (benzoyloxycarbonyl-Asp-CH2OC(O)-2,6-dichlorobenzene)-sensit ive, caspase-3 like cysteine protease that cleaved poly (ADP-ribose) polymerase (PARP), U1 small nuclear ribonucleoprotein (U1 snRNP), and the fluorogenic substrate N-acetyl-Asp-Glu-Val-Asp-7-amido-4-methylcoumarin. In association with these early apoptotic alterations p21 (WAF1/Cip1) is upregulated, but NO affected cell proliferation and apoptosis at a similar dose. At later time points the classical antiapoptotic protein Bcl-2 is downregulated, indicating that decreased Bcl-2 expression is secondary and not a prerequisite for initiation of apoptosis. N-Acetylcysteine (1 mM) interfered with NO-mediated apoptotic signaling, blocking DNA fragmentation as well as PARP and U1 snRNP cleavage. In contrast Z-A-DCB suppressed DNA fragmentation and U1 snRNP cleavage, while PARP breakdown proceeded unaltered. Observing proteolytic PARP digestion without apoptotic alterations questions PARP cleavage as an apoptotic parameter. These results suggest that a Z-A-DCB-sensitive caspase that is distinct from the PARP-cleaving enzyme is activated during NO exposure. NO-mediated apoptotic signaling in U937 cells activates caspases, some of which are dispensable for propagating the death signal.
Brockhaus
F
F
Faculty of Medicine, Department of Medicine IV-Experimental Division, University of Erlangen-Nürnberg, Germany.
Brüne
B
B
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D053938:DNA Fragmentation; D004273:DNA, Neoplasm / Q000032:analysis; D004789:Enzyme Activation; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005978:Glutathione / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D009569:Nitric Oxide / Q000494:pharmacology*; D009589:Nitrogen Oxides; D009603:Nitroso Compounds / Q000494:pharmacology*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis*; D017411:Ribonucleoproteins, Small Nuclear / Q000378:metabolism; D026422:S-Nitrosoglutathione; D013096:Spermine / Q000031:analogs & derivatives* / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D009589:Nitrogen Oxides; D009603:Nitroso Compounds; D019253:Proto-Oncogene Proteins c-bcl-2; D017411:Ribonucleoproteins, Small Nuclear; C091861:spermine nitric oxide complex; D013096:Spermine; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D003546:Cysteine Endopeptidases; D005978:Glutathione